A Whole Blood Bactericidal Assay for Tuberculosis

Wallis, Robert S.; Palaci, Moisés; Vinhas, Solange Alves; Hise, Amy G.; Ribeiro, Fabíola C.; Landen, Katherine; Cheon, Seon Hee; Song, Ho‐Yeon; Phillips, Murray G.; Dietze, Reynaldo; Ellner, Jerrold J.

doi:10.1086/319679

Cited by 95 publications

(115 citation statements)

References 12 publications

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“…Several attempts have been made to bridge the gap between simple ex vivo (macrophage) and in vivo (animal model) experiments, and two examples are cited. A whole-blood assay has been used to study early stages in M. tuberculosis infection, and it utilizes blood drawn from TB patients or noninfected individuals (303). The advantages of this system are that monocytes and lymphocytes present in blood can interact as they do ordinarily and that M. tuberculosis is rapidly phagocytosed and remains intracellular for at leat 72 h. Another system assembles cellular components thought to be involved in early M. tuberculosis infection, by using a tissue culture bilayer system (28).…”

Section: New Ways To Study M Tuberculosis-host Interactionsmentioning

confidence: 99%

Mycobacterium tuberculosisPathogenesis and Molecular Determinants of Virulence

2003

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Tuberculosis (TB), one of the oldest known human diseases. is still is one of the major causes of mortality, since two million people die each year from this malady. TB has many manifestations, affecting bone, the central nervous system, and many other organ systems, but it is primarily a pulmonary disease that is initiated by the deposition of Mycobacterium tuberculosis, contained in aerosol droplets, onto lung alveolar surfaces. From this point, the progression of the disease can have several outcomes, determined largely by the response of the host immune system. The efficacy of this response is affected by intrinsic factors such as the genetics of the immune system as well as extrinsic factors, e.g., insults to the immune system and the nutritional and physiological state of the host. In addition, the pathogen may play a role in disease progression since some M. tuberculosis strains are reportedly more virulent than others, as defined by increased transmissibility as well as being associated with higher morbidity and mortality in infected individuals. Despite the widespread use of an attenuated live vaccine and several antibiotics, there is more TB than ever before, requiring new vaccines and drugs and more specific and rapid diagnostics. Researchers are utilizing information obtained from the complete sequence of the M. tuberculosis genome and from new genetic and physiological methods to identify targets in M. tuberculosis that will aid in the development of these sorely needed antitubercular agents

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Section: New Ways To Study M Tuberculosis-host Interactionsmentioning

confidence: 99%

Mycobacterium tuberculosisPathogenesis and Molecular Determinants of Virulence

2003

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“…Other immune cells, like lymphocytes, have been described in cocultures with infected macrophages. 42 Finally, a very interesting model is whole blood 47 , which contains many components of the immune system that may cooperate in the elimination of the bacilli but is, on the other hand, very difficult to analyze given the complexity of the cellular and humoral interactions that take place in this tissue.…”

Section: Human Cellular Modelsmentioning

confidence: 99%

“…The first one was the [H 3 ]-thymidine incorporation to the nucleic acids from intracellularly growing mycobacteria. 67 Later, other techniques were devised, including quantification by use of the BACTEC system (Becton Dickinson) 47 or by using a mycobacterial reporter strain (usually BCG) transformed with luc, that encodes the enzyme luciferase. 68 Nevertheless, determination of CFU remains the method of choice for most researchers.…”

Section: Cell Lysis and Quantification Of Mycobacterial Growthmentioning

confidence: 99%

In vitro infection of human cells with Mycobacterium tuberculosis

Rivero-Lezcano

2013

Tuberculosis

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It has been estimated that approximately 50% of individuals exposed to Mycobacterium tuberculosis never become tuberculin skin test positive, which may indicate that successful human immunological responses are able not only to inhibit mycobacterial growth, but also to kill the bacteria. Nevertheless, it has been extremely difficult to reproduce this effect in vitro and the ability of human phagocytes to eliminate the bacteria is controversial. This is one of the reasons why we do not fully understand either tuberculosis resistance or susceptibility. Nowadays there is a pressing

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“…1). We and others have previously used whole blood cultures to study acquired host immune responses following vaccination with Mycobacterium bovis BCG (1,4,16). The model's simplicity permits the study of relatively large numbers of strains and subjects.…”

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confidence: 99%

“…Care was taken to ensure that the recent clinical isolates were minimally passaged prior to study to avoid the selection of substrains that were adapted to in vitro replication. The reference strains, which had undergone extensive in vitro culture in broth, included HN878 (also known as 210), a group W-Beijing, outbreak-associated strain; CDC1551, a strain associated with an unusually large number of tuberculin skin test conversions; H 37 Rv, a widely studied reference isolate that is virulent in animal models; H 37 Ra, H 37 Rv's highly attenuated sibling; and MP-28, an isolate that was selected at random from a similar study conducted in Brazil in 1997 (18) and that was cultured extensively in vitro during the course of three subsequent whole-blood infection studies, in which it was identified as strain 28 (1,16,19). The objective of the present project was to examine the influence of strain variation in M. tuberculosis on the early events of human tuberculosis (TB) pathogenesis.…”

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confidence: 99%

Survival and Replication of Clinical Mycobacterium tuberculosis Isolates in the Context of Human Innate Immunity

et al. 2005

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The initial host response to Mycobacterium tuberculosis is driven by innate immunity. For this study, we examined the ability of 18 recent clinical isolates and 5 reference strains to survive and replicate in the context of host innate immunity by using whole blood culture. Six healthy tuberculin-negative volunteers served as subjects. H 37 Ra showed the least capacity to replicate of any of the strains tested, decreasing in viability 1.3 log CFU during 72 h of whole blood culture, whereas H 37 Rv increased 0.32 log. Clinical isolates varied greatly in their ability to replicate in blood cells, ranging from ؊0.4 to ؉0.8 log (P < 0.001). Four showed significantly more growth than H 37 Rv, and one showed significantly reduced growth. Host mechanisms for restricting intracellular mycobacterial growth were more effective during the first 24 h of whole blood culture than during the 24-to 72-h period. Certain mycobacterial isolates appeared preferentially able to withstand host defenses during each of these intervals. Although there was relatively more homogeneity among subjects than among strains, one of the six subjects showed a reduced capacity to restrict intracellular mycobacterial growth due to a defect expressed during the first 24 h of culture. Our findings indicate substantial variability in the capacity of clinical tuberculosis isolates to replicate in host cells in the face of innate host immunity.The early events following inhalation by an immunocompetent, mycobacterium-naïve host of droplet nuclei containing viable Mycobacterium tuberculosis are driven by the innate immune system. The resulting influx of neutrophils, macrophages, NK cells, and other cells to the site of infection serves as a stimulus for granuloma formation and acts directly to limit the extent of mycobacterial replication at this early stage of infection. It has been suggested that the efficiency of these early innate responses may help determine whether a latent infection is established and whether that infection ultimately will progress to active disease.Through evolutionary selection, pathogenic mycobacteria have acquired means to evade specific host immune effector mechanisms, presumably including those of the innate response. The propensity of certain M. tuberculosis isolates to cause outbreaks, for example, has been linked to increased virulence in macrophages or mice in association with altered host cytokine expression profiles (2,3,6,8,12). However, even these virulent outbreak-associated isolates cause disease in only a small proportion of infected individuals. Our understanding of the interplay of biologic and genetic diversity in the host and in the mycobacterium is incomplete, in part because current models to examine the early events in mycobacterial pathogenesis have not been well suited to field or epidemiologic human studies.For the present study, we assessed the capacity of mycobacterial strains to survive phagocytosis and replicate in whole blood cultures. This model permits the involvement of neutrophils, as well as mo...

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A Whole Blood Bactericidal Assay for Tuberculosis

Cited by 95 publications

References 12 publications

Mycobacterium tuberculosisPathogenesis and Molecular Determinants of Virulence

Mycobacterium tuberculosisPathogenesis and Molecular Determinants of Virulence

In vitro infection of human cells with Mycobacterium tuberculosis

Survival and Replication of Clinical Mycobacterium tuberculosis Isolates in the Context of Human Innate Immunity

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